Non-natural MIC proteins
This invention describes soluble, monovalent, non-natural protein molecules that can activate NK cells and certain T-cells to attack specific cellular target cells by attaching the NKG2D-binding portions of monovalent MICA or MICB protein, i.e. their α1-α2 platform domain, to the intended target cell specifically. The α1-α2 domain is contiguous with a heterologous α3 domain that has been genetically modified to bind directly or indirectly to the extracellular aspect of the target cell, thereby serving as the targeting domain. The genetic modification to create a non-natural and non-terminal targeting motif within the α3 domain can include a portion of an antibody, another protein molecule or portion thereof, a peptide, or a non-natural, modified α3 domain of a MIC protein.
1. A non-natural, monomeric, soluble, mammalian MHC class I chain-related (MIC) molecule comprising an α1-α2 platform domain attached to a targeting motif,
wherein the α1-α2 platform domain is at least 80% identical to a native α1-α2 platform domain of a human MICA or MICB protein, and wherein the α1-α2 platform domain binds an NKG2D receptor, and
wherein the targeting motif comprises a MIC α3 domain and a heterologous peptide, wherein the heterologous peptide is inserted into the MIC α3 domain at a non-carboxy-terminal site, and wherein the heterologous peptide directs the binding of the targeting motif to a target molecule on a target cell, thereby delivering the attached α1-α2 platform domain to the target cell.
2. The molecule of claim 1 wherein the α3 domain is from a human MICA or MICB protein.
3. The molecule of claim 1 wherein the α3 domain is a complete native α3 domain without a deletion.
4. The molecule of claim 1 wherein the α3 domain is a native α3 domain, wherein a portion of the domain has been deleted.
5. The molecule of claim 4 , wherein the portion deleted is adjacent to the insertion site.
6. The molecule of claim 1 wherein the α3 domain comprises a deletion, insertion, amino acid substitution, mutation, or combination thereof at site different from the insertion site.
7. The molecule of claim 1 wherein the MICA protein is selected from the group consisting of SEQ ID NOs:1-6, and 13.
8. The molecule of claim 1 , wherein the target molecule is a cell-surface molecule.
9. The molecule of claim 1 wherein the target cell is malignant and the target molecule is a human epidermal growth factor receptor 2 (HER2), NK-1R, Epidermal Growth Factor Receptor (EGFR), Erb2 or melanoma antigen; antigens of LNcaP and PC-3 prostate cancer cells; a growth factor receptor, an angiogenic factor receptor, an integrin, or an oncogene-encoded protein product, or a fragment thereof.
10. The molecule of claim 1 wherein the target cell is infected by a virus and the target molecule on the target cell is a phosphotidylserine, or a phosphotidylserine with an accessory protein; or a surface glycoprotein encoded by a virus, an adenovirus, a human immunodeficiency virus, a herpetic virus, a pox virus, a flavivirus, a filovirus, a hepatitis virus, a papilloma virus, cytomegalovirus, vaccinia, rotavirus, influenza, a parvo virus, West Nile virus, rabies, polyoma, rubella, distemper virus, or Japanese encephalitis virus.
11. The molecule of claim 1 wherein the heterologous peptide is a complementarity determining region of an antibody.
12. The molecule of claim 1 wherein the insertion is within or adjacent to a solvent-exposed loop of the α3 domain.
13. A composition comprising the non-natural, monomeric, MIC molecule of claim 1 and a carrier or excipient.